Elucidation of the mechanism enhancing the avidity of lectin with oligosaccharides on the solid phase surface.
The mechanism underlying molecular recognition of lectins was elucidated by a novel solid phase binding assay system based on surface plasmon resonance. When the apparent affinities of interactions between chitooligosaccharides and wheat germ agglutinin were compared between lectin-immobilized and oligosaccharide-immobilized assay systems, the affinity constants (Ka) calculated for the former system were in good agreement with the previously reported values measured in solution. On the other hand, in the latter system, the calculated Ka could be more than 10,000 times higher than the values in solution at lower lectin concentrations. To elucidate the reason for this, we systematically investigated the effects of the oligosaccharide immobilized density and the lectin valence on the apparent affinity in the oligosaccharide-immobilized assay system. Both the apparent association (k[ass]) and dissociation rate constants (k[diss]) showed a tendency to decrease as the oligosaccharide density increased. This effect was most remarkable for the interaction possessing an extremely fast intrinsic k(ass). Oligomerization of lectin enhanced the avidity due to a significant reduction in k(diss). These phenomena could be explained by considering the nonhomogeneous conditions under which binding occurred. The reaction in a nonhomogeneous state is limited by the mass transport effect, and the effect of rebinding becomes so large that it cannot be disregarded. These findings are the first to demonstrate the importance of the mass transport effect in modulating the affinity of lectin for oligosaccharides on a solid phase surface.
Shinohara
Y
Y
Department of R&D, Pharmacia Biotech K.K, Tokyo, Japan.
Hasegawa
Y
Y
Kaku
H
H
Shibuya
N
N
D000606:Amino Sugars / Q000737:chemistry / Q000378:metabolism; D015374:Biosensing Techniques; D019921:Biotinylation; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002686:Chitin / Q000737:chemistry; D004187:Disaccharides / Q000737:chemistry; D007700:Kinetics; D037102:Lectins / Q000737:chemistry / Q000378:metabolism*; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism*; D010835:Phytohemagglutinins / Q000737:chemistry / Q000378:metabolism; D037121:Plant Lectins; D054788:Ribosome Inactivating Proteins; D014312:Trisaccharides / Q000737:chemistry; D014909:Wheat Germ Agglutinins / Q000737:chemistry / Q000378:metabolism
D000606:Amino Sugars; D004187:Disaccharides; D037102:Lectins; D009844:Oligosaccharides; D010835:Phytohemagglutinins; D037121:Plant Lectins; C017707:Ricinus communis agglutinin-1; C050600:Sambucus nigra lectins; D014312:Trisaccharides; D014909:Wheat Germ Agglutinins; C100156:leukoagglutinins, plants; D002686:Chitin; C000458:N-acetyllactosamine; C041161:chitotriose; C012238:chitotetrose; C032438:chitobiose; D054788:Ribosome Inactivating Proteins